Robustness of Single-cell RNA-Seq for Identifying Differentially Expressed Genes
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE226163 | GEO | 2023/02/27
REPOSITORIES: GEO
Similar Datasets
Project description:Recent advances in RNA sequencing (RNA-Seq) have enabled the discovery of novel transcriptomic variations that are not possible with traditional microarray-based methods. Tissue and cell specific transcriptome changes during pathophysiological stress, in disease cases versus controls and in response to therapies are of particular interest to investigators studying cardiometabolic diseases. Thus, knowledge on the relationships between sequencing depth and detection of transcriptomic variation is needed for designing RNA-Seq experiments and for interpreting results of analyses. Using deeply sequenced RNA-Seq data derived from adipose of a healthy individual before and after systemic administration of endotoxin (LPS), we investigated the sequencing depths needed for studies of gene expression and alternative splicing (AS). We found that to detect expressed genes and AS events, ~100 million (M) filtered reads were needed. However, the requirement on sequencing depth for the detection of LPS modulated differential expression (DE) and differential alternative splicing (DAS) was much higher. To detect 80% of events, ~300M filtered reads were needed for DE analysis whereas at least 400M filtered reads were necessary for detecting DAS. Although the majority of expressed genes and AS events can be detected with modest sequencing depths (~100M filtered reads), the estimated gene expression levels and exon/intron inclusion levels were less accurate. We report the first study that evaluates the relationship between RNA-Seq depth and the ability to detect DE and DAS in human adipose. Our results suggest that a much higher sequencing depth is needed to reliably identify DAS events than for DE genes. Random sampling the RNA-seq data in different depth for gene and alternative-splicing analysis
2013-07-12 | E-GEOD-46323 | biostudies-arrayexpress
Project description:Recent advances in RNA sequencing (RNA-Seq) have enabled the discovery of novel transcriptomic variations that are not possible with traditional microarray-based methods. Tissue and cell specific transcriptome changes during pathophysiological stress, in disease cases versus controls and in response to therapies are of particular interest to investigators studying cardiometabolic diseases. Thus, knowledge on the relationships between sequencing depth and detection of transcriptomic variation is needed for designing RNA-Seq experiments and for interpreting results of analyses. Using deeply sequenced RNA-Seq data derived from adipose of a healthy individual before and after systemic administration of endotoxin (LPS), we investigated the sequencing depths needed for studies of gene expression and alternative splicing (AS). We found that to detect expressed genes and AS events, ~100 million (M) filtered reads were needed. However, the requirement on sequencing depth for the detection of LPS modulated differential expression (DE) and differential alternative splicing (DAS) was much higher. To detect 80% of events, ~300M filtered reads were needed for DE analysis whereas at least 400M filtered reads were necessary for detecting DAS. Although the majority of expressed genes and AS events can be detected with modest sequencing depths (~100M filtered reads), the estimated gene expression levels and exon/intron inclusion levels were less accurate. We report the first study that evaluates the relationship between RNA-Seq depth and the ability to detect DE and DAS in human adipose. Our results suggest that a much higher sequencing depth is needed to reliably identify DAS events than for DE genes.
2013-07-12 | GSE46323 | GEO
Project description:Modest transcriptional changes has been previously observed in solid tumor cell lines in response to inhibition of SUMOylation by ML-792 and here confirmed with TAK-981, lacking a robust induction of IFN1 genes. Comparable modulation of gene expression in HCT116, MDA-MB-231 and Colo-205 cells by TAK-981 and ML-792. Biological triplicates of HCT116, MDA-MB-231 and Colo-205 cells treated with DMSO, 1 μM ML-792 or 1 μM TAK-981 in vitro for 16 h were analyzed by RNA-Seq. The number of differentially expressed genes (DEGs) with false discovery rate corrected p- values of <0.05 and fold-change values greater than 2 are captured. The degree of gene regulation by TAK-981 is plotted against the degree of gene regulation by ML-792 for the union of their DEGs for each cell line. Data are fitted by linear regression after outliers were removed (7 DEGs from Colo- 205, r^2 = 0.951; 2 DEGs from HCT116, r^2 = 0.836; and 2 DEGs from MDA-MB-231, r^2 = 0.971). Limited and comparable upregulation of human IFN1 mRNA signature in Colo-205, and MDA-MB-231 cells, but not in HCT116 cells, following 16h of treatment with 1uM ML-792 or 1uM TAK-981, as determined by human IFN1 ssGSEA gene signature scores. In vitro treatment of the A20, B16F10, CT26 and MC38 mouse tumor cell lines with TAK-981 did not result in induction of the IFN1 pathway. The overall degree of transcriptional modulation by TAK-981 was relatively modest in the mouse tumor cell lines, with only limited overlap observed in the transcriptional responses to TAK-981, as also observed for the human solid tumor cell lines.
2021-04-23 | GSE173116 | GEO
Project description:Sequencing of 46 maize leaf whole transcriptome revealed 77 differentially expressed genes (DEGs) between Lancaster and Non-Lancaster maize genetic resources under optimal growing conditions. Cold test of the subset of four Lancaster and four Non-Lancaster lines showed that the first were cold sensitive and the later cold tolerant, with the majority of identified DEGs showing FPKM values above the plate mean in Lancaster, but below the plate mean in Non-Lancaster subset. Cold induced expression analysis for revealed that, among seven tested DEGs, ATP-sulfurylase and photosystem II I encoding genes showed responsiveness to low temperatures in a genetic-background-dependent manner, likely contributing to different cold response/adaptability of Lancaster and Non-Lancaster lines.
2020-12-31 | GSE164078 | GEO
Project description:Many critical infrastructure systems have network structures and are under stress. Despite their national importance, the complexity of large-scale transport networks means that we do not fully understand their vulnerabilities to cascade failures. The research conducted through this paper examines the interdependent rail networks in Greater London and surrounding commuter area. We focus on the morning commuter hours, where the system is under the most demand stress. There is increasing evidence that the topological shape of the network plays an important role in dynamic cascades. Here, we examine whether the different topological measures of resilience (stability) or robustness (failure) are more appropriate for understanding poor railway performance. The results show that resilience, not robustness, has a strong correlation with the consumer experience statistics. Our results are a way of describing the complexity of cascade dynamics on networks without the involvement of detailed agent-based models, showing that cascade effects are more responsible for poor performance than failures. The network science analysis hints at pathways towards making the network structure more resilient by reducing feedback loops.
| S-EPMC6408419 | biostudies-literature
Project description:Mesenchymal stem cells (MSCs) possess the potential of differentiating into chondrocytes, which makes it an ideal source in healing cartilage defects. Here, we want to find out the key genes that participate in MSCs chondrogenesis. Human MSCs were induced for chondrogenesis for 7, 14, and 21 days using high-density micromass culture system and RNA was extracted for RNA-seq. A total of 6247 differentially expressed gene (DEGs) were identified at day 7, and 85 DEGs were identified at day 14. However, no significant DEGs was identified in day 21. The top 30 DEGs at day 7, including COL9A3, COL10A1, and CILP2, are closely related to extracellular matrix organization. While the top 30 DEGs at day 14 revealed that genes related to inflammation were enriched, including CXCL8, TLR2, and CCL20. We also conducted protein-protein interaction (PPI) networks analysis using search tool for the retrieval of interacting genes (STRING) database and identified key hub genes including CXCL8, TLR2, CCL20, and MMP3. The transcriptional factors were also analyzed, and identified top 5 TFs: LEF1, FOXO1, RORA, BHLHE41, SOX5. We demonstrated one particular TF, RORA, in promoting early MSCs chondrogenesis. Taken together, our result suggested that these DEGs may have a complex effect on MSCs chondrogenesis both synergistically and solitarily.
2022-08-13 | GSE210984 | GEO
Project description:Transcriptome analysis revealed that 23 differentially expressed genes (DEGs) were upregulated and 17 DEGs were downregulated in SKOV3 cells. In OVCAR3 cells, 198 DEGs were upregulated, and 17 DEGs were downregulated. Notably, 53 DEGs exhibited statistically significant differences among the 1,612 DEGs identified. Our findings indicate that HERV-K env gene KO exerts a profound influence on gene expression patterns in OVCAR3 cells, while genetic alterations in expression were relatively modest in SKOV3 cells. Nevertheless, genes ND1, ND2, and CYTB displayed a common increase in expression, while ERRFI1 and NDRG1 exhibited a decrease in expression in both cell lines.
2024-10-31 | GSE269969 | GEO
Project description:In a previous study, we found that H2S alleviates salinity stress in cucumber by maintaining the Na+/K+ balance and by regulating H2S metabolism and the oxidative stress response. However, little is known about the molecular mechanisms behind H2S-regulated salt-stress tolerance in cucumber. Here, an integrated transcriptomic and proteomic analysis based on RNA-seq and 2-DE was used to investigate the global mechanism underlying H2S-regulated salt-stress tolerance. In total, 11 761 differentially expressed genes (DEGs) and 61 differentially expressed proteins (DEPs) were identified. Analysis of the pathways associated with the DEGs showed that salt stress enriched expression of genes in primary and energy metabolism, such as photosynthesis, carbon metabolism and biosynthesis of amino acids. Application of H2S significantly decreased these DEGs but enriched DEGs related to plant-pathogen interaction, sulfur-containing metabolism, cell defense and signal transduction pathways. Notably, changes related to sulfur-containing metabolism and cell defense were also observed through proteome analysis, such as Cysteine synthase 1, Glutathione S-transferase U25-like, Protein disulfide-isomerase and Peroxidase 2. We present the first global analysis of the mechanism underlying H2S regulation of salt-stress tolerance in cucumber through tracking changes in the expression of specific proteins and genes.
2021-09-09 | PXD020642 | Pride
Project description:The dynamic behavior of metabolic networks is governed by numerous regulatory mechanisms, such as reversible phosphorylation, binding of allosteric effectors or temporal gene expression, by which the activity of the participating enzymes can be adjusted to the functional requirements of the cell. For most of the cellular enzymes, such regulatory mechanisms are at best qualitatively known, whereas detailed enzyme-kinetic models are lacking. To explore the possible dynamic behavior of metabolic networks in cases of lacking or incomplete enzyme-kinetic information, we present a computational approach based on structural kinetic modeling. We derive statistical measures for the relative impact of enzyme-kinetic parameters on dynamic properties (such as local stability) and apply our approach to the metabolism of human erythrocytes. Our findings show that allosteric enzyme regulation significantly enhances the stability of the network and extends its potential dynamic behavior. Moreover, our approach allows to differentiate quantitatively between metabolic states related to senescence and metabolic collapse of the human erythrocyte. We think that the proposed method represents an important intermediate step on the long way from topological network analysis to detailed kinetic modeling of complex metabolic networks.
| S-EPMC2132447 | biostudies-other
Project description:The "reproducibility crisis" in science affects microbiology as much as any other area of inquiry, and microbiologists have long struggled to make their research reproducible. We need to respect that ensuring that our methods and results are sufficiently transparent is difficult. This difficulty is compounded in interdisciplinary fields such as microbiome research. There are many reasons why a researcher is unable to reproduce a previous result, and even if a result is reproducible, it may not be correct. Furthermore, failures to reproduce previous results have much to teach us about the scientific process and microbial life itself. This Perspective delineates a framework for identifying and overcoming threats to reproducibility, replicability, robustness, and generalizability of microbiome research. Instead of seeing signs of a crisis in others' work, we need to appreciate the technical and social difficulties that limit reproducibility in the work of others as well as our own.
| S-EPMC5989067 | biostudies-literature